This project concerns mechanisms of CNS demyelination in human disease and continues to focus on the human JC polyomavirus (JCV), the etiologic agent of progressive multifocal leukoencephalopathy (PML). PML is a frequently fatal demyelinating disease which complicates up to 5% of AIDS cases, and can signal onset of clinical AIDS. Work this year has emphasized the detection of the JC virus and BK virus (BKV) by polymerase chain reaction (PCR) in PML tissues, normal human brain tissues, human kidneys, urine, Kaposi~s sarcoma, and infected hamster tissues. Notable advances have concerned the use of reverse transcription PCR (RT~PCR) to detect JCV RNA, evidence that rearrangement of the JCV regulatory region occurs within the host prior to or during onset of PML, identification of a mutation in BKV apparently leading to tubulo~interstitial nephritis and end~stage renal disease in an AIDS patient, and the detection of JCV in the urine and kidneys from multiple sclerosis (MS) patients (parts of this work are also reported under related projects from the Neurotoxicology Section). The methods for typing JCV coding regions using type~specific primers previously developed in this Section have been applied to PML cases from Japan and Germany originally reported as due to SV40. Instead, JCV Type 1 was found in the German case, and JCV Type 2 in the Japanese case. Typing on paraffin~embedded tissues from an additional 33 PML cases is underway. We have sought to confirm previous reports from this Section and elsewhere that JCV sequences are present in some normal human brains. We have also examined MS brain sections for JCV DNA. To date, the results from both normal and MS brains have been negative with either ordinary PCR, nested PCR, or with intron~differential RNA PCR. However, approximately 40% of urines from MS patients were found to contain JCV DNA, and 2 of 9 MS kidneys also harbored JCV DNA. Further characterization of these JCV~MS strains is under way to determine whether they differ significant ly from previously characterized JCV~PML strains. A kidney from a case of tubulo~interstitial nephritis with end~stage renal disease in an AIDS patient has been shown to harbor both JCV and BKV. The BKV DNA amplified from this kidney was mutated in a region of the large T~antigen gene though to be involved in DNA binding based on previous reports from studies with SV40. This finding raises the specter that more dangerous forms of usually harmless DNA viruses may emerge in the context of human immunodeficiency virus type 1 (HIV~1) infection.